The goal of this proposal is to understand the role of the Sen1 helicase in termination of RNA Polymerase II (PolII) transcripts and to establish how mutations in the human homolog of Sen1, Senataxin (SETX), cause the neurodegenerative disorders Ataxia-Oculomotor-Apraxia type 2 (AOA2) and Amyotrophic Lateral Sclerosis Type 4 (ALS4). Based on the yeast model, we propose that SETX mutations cause human disease through disruption of transcriptional termination. Thus, we want to define the precise function of the Sen1/SETX helicases in transcriptional termination and the manner in which these functions are disrupted in disease. The first aim of the grant is to learn more about the role of yeast Sen1 in transcription termination and its regulation by other components of the transcription termination machinery. We will determine whether Sen1 terminates PolII by direct action, or indirectly via its effect on small nucleolar ribonucleoprotein (snoRNP) assembly. We will accomplish this via analysis of the protein and RNA composition of the snoRNP particles under suboptimal cellular Sen1 levels. Next, we will identify the mechanism of the cell death caused by mutations of Sen1. This will be accomplished through alleviating the promoter occlusion effect of snoRNA messages reading into downstream essential genes. If the currently accepted hypothesis is correct, this should suppress the lethality associated with Sen1 loss. Subsequently we will explore the role of phosphorylation in the regulation of Sen1. The second aim is to identify the shared characteristics of yeast Sen1 and human SETX function. We will affinity purify SETX and identify the proteins and transcripts that associate with it. We will also explore how these associations are regulated by phosphorylation. The third aim is to identify the mechanism by which mutations in human SETX cause disease. We will define how mutations in SETX change its protein and RNA binding properties and examine whether mutations in proteins that SETX associates with cause similar defects in cellular metabolism as mutations in SETX. We believe that accomplishment of the goals outlined above will lead to a better understanding of the regulation of transcriptional termination and the pathogenesis of neurodegenerative disorders. The information obtained about the function of SETX might allow us to devise new strategies to reverse the progression of neurodegenerative diseases like ALS4 and AOA2. Project Narrative Our work will investigate how mutations in the Senataxin protein, SETX, cause the neurodegenerative disorders Ataxia Oculomotor Apraxia type 2 (AOA2) and Amyotrophic Lateral Sclerosis Type 4 (ALS4). The knowledge gained might provide new strategies to reverse the progression of neurodegenerative diseases.